ABSTRACT A very dense breast is one of the strongest risk factors for breast cancer. Close to 2 million women aged 50-64 years have extremely dense breast. It is estimated that 28% of breast cancer cases are attributable to increased breast density. Compared with other breast cancer risk factors that confer similar magnitude of risk (age, atypia on breast biopsy and highly penetrant breast cancer susceptibility genes), breast density is modifiable. A decrease in breast density over time is associated with decreased risk of breast cancer. Hence, strategies to reduce breast density could present a path to targeted prevention of breast cancer in many mid- life women. However, there is very limited knowledge on how to modify breast density or biomarkers that can be targeted in reducing breast density. Recent seminal discoveries have identified that receptor activator of nuclear factor-?B (RANK) pathway plays a crucial role in the development of a functional lactating mammary gland as well as regulating mammary epithelial cells. Further, RANK ligand (RANKL) signaling is a major mediator of progesterone-induced proliferation of mammary epithelial cells and disruption of RANKL signaling attenuates progestin-driven mammary epithelial cell proliferation. Therefore, RANKL inhibition could provide a path to reducing breast density and breast cancer incidence in women with increased breast density. A well-tolerated RANKL inhibitor (denosumab) is already in clinical use for treating postmenopausal osteoporosis, but this has not been applied in primary prevention of breast cancer. Its ability to effectively inhibit RANKL signaling could allow for a novel approach to breast cancer prevention. In this proposal, we will investigate the association of the RANK pathway (circulating levels and genetic polymorphisms) with percent mammographic density in mid- life women (50-64). We will also investigate factors that influence these women's intent to use a new chemopreventive agent. Study participants (N=398) will be recruited among women undergoing annual screening mammogram at the Breast Health Center (BHC), Washington University School of Medicine (WUSM), St. Louis. Blood draw will take place on the day of screening mammogram. A detailed reproductive and lifestyle questionnaire will also be completed. We will use Volpara software to assess percent breast density. Volpara permits robust quantitative analysis of breast density. Our proposal is innovative and could demonstrate novel, functionally important mechanistic associations with mammographic density. Importantly, it could provide essential information for the launch of trials targeting RANKL signaling in the primary prevention of breast cancer among mid-life women with very dense breast. 1